Host responses to recombinant hemagglutinin B of Porphyromonas gingivalis in an experimental rat model

Porphyromonas gingivalis, a gram-negative, black-pigmented anaerobe, is amo ng the microorganisms implicated in the etiology of adult periodontal disea se, This bacterium possesses a number of factors, including hemagglutinins, of potential importance in virulence. Several hemagglutinin genes have bee n identified, cloned, and expressed in Escherichia coli, The purpose of thi s study was to characterize host responses to purified recombinant hemagglu tinin B (rHag B), using the conventional Fischer rat as the experimental an imal model. The effectiveness of immunization with rHag B on protection aga inst experimental periodontal bone loss following infection with P. gingiva lis was also evaluated, Groups of rats were immunized by the subcutaneous r oute with rHag B in complete Freund's adjuvant, immunized with rHag B and o rally infected with P. gingivalis, nonimmunized and noninfected, or orally infected with P. gingivalis only. Serum and saliva samples were collected t hroughout the experiment and evaluated for serum immunoglobulin G (IgG) and IgM and salivary IgA antibody activity by enzyme-linked immunosorbent assa y. No salivary IgA anti-Nag B activity was detected in the various groups o f rats. A slight serum IgM response similar to that seen in preimmune sampl es was observed, Serum IgG antibody activity to Hag B was detected only in samples from rats immunized with rHag B, This response was primarily of the IgG1 and IgG2a subclasses, followed by IgG2b and low levels of IgG2c. Supe rnatants from rHag B-stimulated splenic lymphoid cell cultures from immuniz ed rats contained high levels of gamma interferon, followed by interleukin- 2 (IL-2), IL-10, and then IL-4. These results are consistent with the induc tion of T helper type 1 (Th1)- and Th2-like responses. Western blot analysi s of sera derived from rHag B-immunized rats reacted with trichloroacetic a cid (TCA) precipitates of P. gingivalis 33277, 381, A7A1-28, and W50, revea ling a 50-kDa band reflective of Hag B, However, sera derived from rats imm unized with P. gingivalis whole cells or from rats infected with P, gingiva lis only did not react with rHag B but did react with TCA precipitates of P , gingivalis strains. Finally, radiographic measurements of periodontal bon e loss indicated that rats immunized with rHag B had less bone loss than th ose infected with P. gingivalis only. These results demonstrate the effecti veness of purified rHag B in inducing a protective immune response and supp ort the potential usefulness of this component of P. gingivalis in the deve lopment of a vaccine against adult periodontitis.